Development and fine structure of murine Purkinje cells in dissociated cerebellar cultures: neuronal polarity.
Cerebellar Purkinje cells (PC) display a highly distinctive form of polarity. We have cultured murine PCs from dissociated E16 cerebellar anlagen for 1 week to investigate the early stages of neuronal compartmentalization and synaptic interactions, features which are important for the establishment of neuronal polarity. To unequivocally identify the PCs we utilized light and electron microscopic immunocytochemistry with an anti-serum to the cell class-specific marker L7/pcp2 gene product. The PCs typically show a single, long axon, numerous short appendages classified as filopodia and protospines, and a small number of protodendrites. The nucleus is positioned asymmetrically in both the horizontal and vertical axes of the soma. The Golgi apparatus, coated and uncoated vesicles, and mitochondria are prominent ultrastructural features, while the endoplasmic reticulum is highly fragmented. The cell body receives rudimentary synapses on its smooth surfaces and appendages and no consistent morphological differences were detected between these elementary contacts. The axon is clearly identifiable; it emanates from either the cell body or a protodendrite, bifurcates at predominantly right angles, forms beaded collaterals, and terminates with relatively large growth cones. The varicosities of the PC axon contain pleomorphic synaptic vesicles and form rudimentary synapses primarily with the dendritic shafts of immunonegative neurons. The protodendrites are short, quickly tapering and sparsely branched; they emit numerous filopodia and immature spines and terminate with small growth cones. Rudimentary synapses are received on the proximal dendritic shafts and filopodia, and more mature synapses occur frequently on protospines. With few exceptions, PCs lie atop an astrocytic bed layer and glial processes are apposed to the various aspects of the PC body left free by the afferent axons. By contrast, PC processes are largely free of glial sheaths. We conclude that the "stellate stage" of PC development in situ is replicated rather faithfully in culture and that PCs have established polarity and have begun to form intercellular contacts by 1 week in vitro. Moreover, the PCs are already morphologically distinct from other cell types in the 1 week cultures, although they have yet to develop the differentiated features that distinguish mature PCs.
Dunn
M E
ME
Department of Neurology Research, Children's Hospital of Philadelphia, PA 19014, USA.
Schilling
K
K
Mugnaini
E
E
D000818:Animals; D001369:Axons / Q000648:ultrastructure; D002450:Cell Communication / Q000502:physiology; D016764:Cell Polarity / Q000502:physiology*; D002478:Cells, Cultured; D002531:Cerebellum / Q000166:cytology* / Q000196:embryology; D003712:Dendrites / Q000648:ultrastructure; D051379:Mice / Q000196:embryology; D008809:Mice, Inbred C3H; D008810:Mice, Inbred C57BL; D009457:Neuroglia / Q000502:physiology; D009474:Neurons / Q000502:physiology; D011689:Purkinje Cells / Q000502:physiology* / Q000648:ultrastructure*; D013569:Synapses / Q000648:ultrastructure